Dipeptide derivatives and their production

ABSTRACT

Dipeptide derivatives represented by the formula: ##STR1## wherein R represents hydrogen, C 1  -C 6  alkyl group, C 2  -C 7  alkenyl group, C 2  -C 7  cyanoalkyl group, C 2  -C 7  carbamoylalkyl group, C 3  -C 10  dialkylaminoalkyl group or cyclopropylmethyl group, R 1  represents hydrogen, C 1  -C 6  alkyl group, C 7  -C 14  aralkyl group, C 7  -C 14  hydroxyaralkyl group, C 6  -C 12  aryl group, C 2  -C 7  carbamoylalkyl group, C 2  -C 7  carboxyalkyl group, C 1  -C 6  aminoalkyl group, C 4  -C 10  guanidylalkyl group, C 1  -C 6  mercaptoalkyl group, C 2  -C 7  alkylthioalkyl group, C 9  -C 15  indolylalkyl group or C 4  -C 9  imidazolylalkyl group, R 2  represents hydrogen, C 1  -C 6  alkyl group, C 7  -C 14  aralkyl group, C 6  -C 12  aryl group, glycyl group or glycyl-glycyl group, R 3  represents hydrogen, C 1  -C 6  alkyl group or amino-protecting group, R 1  and R 2  optionally combine to from C 2  -C 4  alkylene group, group ##STR2## optionally represents phthalimido group, piperidino group, 4-hydroxy-4-(p-halogenophenyl)piperidino, morpholino, or piperazino group substituted by C 1  -C 6  alkyl group or phenyl group, A ring represents benzene ring or pyridine ring optionally substituted by halogen and B ring represents benzene ring or thiophene ring optionally substituted by halogen, trifluoromethyl group, methylsulfonyl group, nitro group or C 1  -C 6  alkyl group and their acid addition salts, being useful as anxiolytics, sedatives, anticonvulsants, hypnotics, muscle relaxants, or their synthetic intermediates, are prepared.

This application is a division of application Ser. No. 601,134, filedAug. 1, 1975 (now abandoned).

The present invention relates to dipeptide derivatives and theirproduction. More particularly, this invention relates to dipeptidederivatives represented by the formula: ##STR3## wherein R representshydrogen, C₁ -C₆ alkyl group, C₂ -C₇ alkenyl group, C₂ -C₇ cyanoalkylgroup, C₂ -C₇ carbamoylalkyl group, C₃ -C₁₀ dialkylaminoalkyl group orcyclopropylmethyl group, R¹ represents hydrogen, C₁ -C₆ alkyl group, C₇-C₁₄ aralkyl group, C₇ -C₁₄ hydroxyaralkyl group, C₆ -C₁₂ aryl group, C₂-C₇ carbamoylalkyl group, C₂ -C₇ carboxyalkyl group, C₁ -C₆ aminoalkylgroup, C₄ -C₁₀ guanidylalkyl group, C₁ -C₆ mercaptoalkyl group, C₂ -C₇alkylthioalkyl group, C₉ -C₁₅ indolylalkyl group or C₄ -C₉imidazolylalkyl group, R² represents hydrogen, C₁ -C₆ alkyl group, C₇-C₁₄ aralkyl group, C₆ -C₁₂ aryl group, glycyl group or glycyl-glycylgroup, R³ represents hydrogen, C₁ -C₆ alkyl group or amino-protectinggroup, R¹ and R² optionally combine to form C₂ -C₄ alkylene group,##STR4## optionally represents phthalimido group, piperidino group,4-hydroxy-4-(p-halogenophenyl)piperidino, morpholino, or piperazinogroup substituted by C₁ -C₆ alkyl group or phenyl group, A ringrepresents benzene ring or pyridine ring optionally substituted byhalogen and B ring represents benzene ring or thiophene ring optionallysubstituted by halogen, trifluoromethyl group, methylsulfonyl group,nitro group or C₁ -C₆ alkyl group and their acid addition salts, beinguseful as anxiolytics, sedatives, anticovulsants, hypnotices, musclerelaxants or their synthetic intermediates.

Illustrative explanation is given to the above definition as follows:alkyl group (e.g. methyl, ethyl, isopropyl, butyl, pentyl), alkenylgroup (e.g. allyl, butenyl, pentadienyl), cyanoalkyl group (e.g.cyanomethyl, cyanoethyl, cyanopropyl, cyanobutyl), carbamoylalkyl group(e.g. carbamoylmethyl, carbamoylethyl, carbamoylpropyl), carboxyalkylgroup (e.g. carboxymethyl, carboxyethyl, carboxypropyl), aminoalkylgroup (e.g. aminomethyl, aminoethyl, aminopropyl, aminobutyl),hydroxyalkyl group (e.g. hydroxymethyl, hydroxyethyl, hydroxypropyl,hydroxybutyl), guanidylalkyl group (e.g. guanidylmethyl, guanidylethyl,guanidylpropyl), mercaptoalkyl group (e.g. mercaptomethyl,mercaptoethyl, mercaptopropyl, mercaptobutyl), alkylthioalkyl group(e.g. methylthiomethyl, ethylthiopropyl, methylthiobutyl), indolylalkylgroup (e.g. indolylmethyl, indolylethyl, indolylpropyl), aralkyl group(e.g. benzyl, phenethyl, phenylpropyl), hydroxyaralkyl group (e.g.hydroxybenzyl, hydroxyphenethyl), aryl group (e.g. phenyl, naphthyl),halogen (e.g. chlorine, bromine, fluorine, iodine), alkylene group (e.g.dimethylene, trimethylene, tetramethylene), amino-protecting group (e.g.carbobenzoxy, methoxycarbonyl, t-butoxycarbonyl,p-methoxybenzyloxycarbonyl, o-nitrophenylsulfenyl,chlorobenzyloxycarbonyl, trityl), and dialkylaminoalkyl group (e.g.dimethylaminoethyl, diethylaminoethyl, diethylaminopropyl).

The dipeptide derivatives (I) can be prepared as shown in the followingscheme: ##STR5## wherein X and X¹ represent each halogen and R, R¹, R²,R³, A ring and B ring each is as defined above but B ring in XI and XIIis benzene ring.

Route A

This route is effected by subjecting the starting amine (II) and theglycine derivative (III) to the amido bonding formation in aconventional manner for the peptide condensation. The amido bondingformation substantially involves the condensation of amino group on theamine (II) with carboxy group on the glycine derivative (III) to formthe peptide bonding and also other accessory treatments for attainingthis object, inclusive of treatment for converting the carboxy group ofthe glycine derivative (III) into its reactive derivatives in advance ofthe amido bonding formation, treatment for previous protection of activegroup (e.g. amino group, carboxy group) which should not participate inthe reaction and treatment for removing such protecting groups after theamido bonding formation. The conversion of the glycine derivative (III)into the reactive derivative involves halogenation, anhydride formation,azide formation, active ester formation, etc. Introduction and removalof such protecting groups can be effected in a conventional manner.Illustrating an example about amino group, the amino group of thepeptide can be protected by treating with carbobenzoxy chloride in thepresence of an alkali, and this amino-protecting group of the finalproduct (I) can be removed by treating with such an acid as hydrobromicacid, hydrofluoric acid or trifluoroacetic acid or by hydrogenating orreducing with liquid ammonia/metallic sodium. Trityl group can beintroduced by treating with trityl chloride in the presence of a baseand can be removed by treating with dilute acetic acid, and phthalylgroup can be removed by treating with hydrazine hydrate. This step isgenerally carried out in an inert solvent (e.g. methylene chloride,dimethylformamide, dimethylsulfoxide, hexamethylphosphoric triamide,chloroform, dioxane, benzene, tetrahydrofuran, a mixture thereof) atroom temperature or under cooling or heating. The general procedure forthe amido bonding formation described in Route A is similarly applicablein the amido bonding formation of other routes hereinafter described.

Route B

This route is effected by subjecting the glycinamide (IV) and the aminoacid (V) to the amido bonding formation. The starting glycylamide beingin a form of its acid addition salts (e.g. hydrobromide, hydrochloride)is also prepared by the amido bonding formation of the amine (II) andglycine. The amido bonding formation of this route is substantiallycarried out as in Route A. For example, the glycinamide (IV) is treatedwith phthalyl-glycyl chloride in a suitable solvent (e.g.dimethylformamide, hexamethylphosphoric triamide) to give thephthalyl-glycyl-glycinamide (I), which is converted into the finalproduct (I) by hydrazinolysis.

Route C

This route is effected in two steps by at first reacting the glycinamide(IV) preferably in a form of its acid addition salt (e.g. hydrochloride,hydrobromide) with the halogenoacetyl halide (VI) to give thehalogenoacetyl-glycinamide (VII) and secondly reacting the latter withthe ammonia, phthalimide or amine (VIII). For elevating the reactivityof the intermediate (VII), the halogen of the compound (VII) may besubstituted by the other more active halogen before the second step, forexample, by treating with alkali halide (e.g. potassium iodide, sodiumiodide, potassium bromide). These reactions are effected in an inertsolvent (e.g. dimethylformamide, hexamethylphosphoric triamide,tetrahydrofuran, acetone, chloroform, diglyme) at room temperature orunder cooling or heating in a conventional manner.

Route D

This route is effected in two steps by subjecting the methylol compound(IX) and the glycine derivative (III) to the amido bonding formation andthen oxidizing the resulting peptide (X). The starting methylol(IX) canbe prepared by reducing the corresponding carbonyl compound (II). Theamido bonding formation is carried out as in Route A, and the oxidationis carried out by treating with such an oxidizing agent as Jones'reagent (chromic acid/sulfuric acid/water), manganese dioxide, chromicanhydride or the like in a conventional manner.

Route E

This route is effected in two steps by subjecting the2-aminomethylindole (XI) and the amino acid (V) to the amido bondingformation and then oxidizing the resulting amide (XII). The amidobonding formation is carried out as in Route A. The oxidation is carriedout by using such an oxidizing agent as oxygen, ozone, hydrogenperoxide, chromic acid, peracid (e.g. peracetic acid), potassiumpermanganese, manganese dioxide or sodium periodate in a conventionalmanner for oxidizing a double bond into carbonyl groups.

When the product (I) contains amino-protecting group, it can be removedaccording to its necessity. Therefor a conventional procedure forremoving amino-protecting group from peptides can be adopted asdescribed in Route A.

The product (I) can be converted into suitable acid addition salts suchas those of inorganic acid (e.g. hydrochloric acid, sulfuric acid,nitric acid, phosphoric acid, thiocyanic acid) or those of organic acid(e.g. acetic acid, succinic acid, oxalic acid, maleic acid, malic acid,phthalic acid, methanesulfonic acid) for the necessity of preparation,crystalization, solubility or improvement of stability.

Thus obtained dipeptide derivatives (I) and their acid addition saltsare useful as anxiolytics, sedatives, anticonvulsives, hypnotics, musclerelaxants, or their synthetic intermediates. Pharmacological activitiesof some dipeptide derivatives (I) are shown in the following table incomparison with chlordiazepoxide and diazepam.

1. Compound tested

    ______________________________________                                        Compound No.                                                                             Compound Name        Note                                          ______________________________________                                        1          2-benzoyl-4-chloro-N-methyl-                                                  N.sup.α -glycyl-glycinanilide                                2          2-o-chlorobenzoyl-4-chloro-N-                                                 methyl-N.sup.α -glycyl-glycinanilide                                    hydrate                                                            3          2-o-fluorobenzoyl-4-chloro-N-                                                 methyl-N.sup.α -glycyl-glycinanilide                                    hydrochloride                                                      4          Chlordiazepoxide     Control                                       5          Diazepam             Control                                       ______________________________________                                    

2. Test method 1. Anti-pentylenetetrazol activity

This test was measured on a group of 10 DS male mice. Within 15 minutesafter subcutaneous injection of 125 mg/kg of pentylenetetrazol, themouse showed tonic convulsion which ceased fatally. In this test, thetest compound was given orally 60 minutes prior to the administration ofpentylenetetrazol. The observation was made for 2 hours after theadministration of pentylenetetrazol. The criterion of anticonvulsantactivity was determined as being complete protection against mortality.The convulsions were disregarded. Results were shown as ED₅₀ [Goodman,et al.: J. Pharmacol., 108, 168 (1953)].

2. Taming activity

This test was measured on DS male mice.

When a 5 Hz square wave pulse (10 msc, 50 v) was delivered to a pair ofmice in a grid box, some pairs showed fighting posture 15 to 20 timesfor 3 minutes. These pairs of mice were selected in the morning and usedfor drug test in the afternoon. The test compound was administered toboth mice of the pairs 60 minutes prior to the experiment. Results wereobtained as the mean percentage of inhibition of fighting responses in agroup of 5 pairs and shown by ED₅₀ [Tedeschi, et al.: J. Pharmacol Exp.Thev., 125, 28-34 (1959)].

3. Rotarod performance activity:

This test was measured on DS male mice.

The mouse was put on a scraped rod of wood, 3 cm in diameter, turning atthe rate of five rotations per minute. The mice that could remain on therod for 3 or more minutes in successive trails were selected and placedin a group of 10 mice for each dose. If the mouse fell down from the rodwithin less than 2 minutes, the test compound was considered to beeffective. Results were shown by ED₅₀ [Dunham, et al.: J. Am. Pharm.Assoc., 46, 208 (1957)].

4. Acute toxicity

The test compound was orally administered to DS male mice in differentsingle doses. For each dose, 10 mice were used, their weight rangingfrom 20 to 23 grams. The mice were observed for 72 hours after theadministration of the compound. The mortality was calculated by theBliss method [Bliss: Ann. Appl. Biol., 22. 134-307 (1935); Qant. J.Pharmacol., 11, 192 (1938)].

3. Result

                  Table 1.                                                        ______________________________________                                                Anti-pentylene-                                                                           Taming   Rotarod  Acute                                           tetrazol acti-                                                                            activity performance                                                                            toxicity                                Compound                                                                              vity, ED.sub.50                                                                           ED.sub.50                                                                              ED.sub.50                                                                              LD.sub.50                               No.     (mg/kg)     (mg/kg)  (mg/kg)  (mg/kg)                                 ______________________________________                                        1       1.0         10.5     36.6     1309                                    2       0.58        3.0      27.7     1255                                    3       0.33        1.6      17.0     >1000                                   4       3.7         24.0     50.0     1090                                    5       1.19        6.0      9.03     1459                                    ______________________________________                                    

4. Conclusion

Each of the five test compounds is very weak in the acute toxicity, andany predominant difference is not observed between them. Compounds onthe subject of this invention (Compound Nos. 1-3) are about 2 to 4 timesless potent in the disturbing effect of motor coordination due to therotarod performance than diazepam (Compound No. 5). In theanti-pentylenetetrazol activity and the taming activity,2-benzoyl-4-chloro-N-methyl-N.sup.α -glycyl-glycinanilide (CompoundNo. 1) is about 2 to 3 times more potent than chlordiazepoxide (CompoundNo. 4), and 2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-glycyl-glycinanilide hydrate (Compound No. 2) and2-o-fluorobenzoyl-4-chloro-N-methyl-N.sup.α -glycyl-glycinanilidehydrochloride (Compound No. 3) are about 2 to 4 times more potent thandiazepam (Compound No. 5).

The dipeptide derivatives (I) and their pharmaceutically acceptable acidaddition salts are applied singly or in combination withpharmaceutically suitable carriers such as wheat starch, corn starch,potato starch, gelatin, etc. The choice of carriers is determined by thepreferred route of administration, the solubility of the substance andstandard pharmaceutical practice. Examples of pharmaceuticalpreparations are tablets, capsules, pills, suspensions, syrups, powders,and solutions. These compositions can be prepared in a conventionalmanner. A suitable dosage of the dipeptide derivatives (I) or theirpharmaceutically acceptable acid addition salts for aduts is in theorder of about 1 mg to 30 mg per day.

Still, the dipeptide derivatives (I) and their acid addition salts areuseful as growth promotors of domestic cattle and fowls.

Presently preferred and practical embodiments of the present inventionare illustratively shown in the following examples.

EXAMPLE 1

1. To a solution of trityl-glycyl-glycine (5 g) in hexamethylphosphorictriamide (24 ml), thionyl chloride (1.6 g) is added dropwise at -8° to-2° C, and the resultant mixture is stirred at -5° C for 20 minutes. Themixture is mixed with 2-amino-5-chlorobenzophenone (3.08 g) and allowedto stand at room temperature overnight. The reaction mixture isneutralized with an aqueous sodium bicarbonate solution and shaken withchloroform. The organic layer is washed with water, dried and evaporatedto remove the solvent. The residue is crystallized from ether to give2-benzoyl-4-chloro-N.sup.α -trityl-glycyl-glycinanilide (1.7 g). Theproduct is recrystallized from ethyl acetate to give needles melting at187° to 189° C. UV: λ_(max) ^(EtOH) 237.5, 274 (sh.), 343 mμ (log ε:4.51, 4.03, 3.53).

2. A suspension of 2-benzoyl-4-chloro-N.sup.α-tritylglycyl-glycinanilide (1.7 g) in 50% acetic acid (20 ml) is heatedon a water bath for 20 minutes. After cooling, the precipitated crystalsare filtered. The filtrate is neutralized with aqueous sodiumbicarbonate solution and shaken with chloroform. The organic layer iswashed with water, dried and evaporated to remove the solvent, whereby2-benzoyl-4-chloro-N.sup.α -glycyl-glycinanilide (0.8 g) is obtained.The product is recrystallized from ethyl acetate to give prisms meltingat 135° to 136° C. UV: λ_(max) ^(EtOH) 241, 275 (sh.), 340 mμ (log ε:4.44, 4.03, 3.55).

EXAMPLES 2-5

Using the following starting materials (II) and (III) the reaction iseffected as in Example 1 to give the corresponding products (Ia) and(Ib): ##STR6## wherein R³ represents amino-protecting group, and R, R¹and R² are as defined above.

                  Table 2.                                                        ______________________________________                                        Example                                                                              II     III            Ia      Ib                                       No.    R      R.sup.1 R.sup.2                                                                            R.sup.3                                                                             mp(° C)                                                                        mp(° C)                       ______________________________________                                        2      H      H       H    Cbz   163-164 135-136                              3       Me    H       H    Tri   Amorph  Amorph                               4      H       Me     H    Cbz   148-149 131-132                              5.sup.a)                                                                             H      i-Bu    H    Cbz    98-100 145-147                              ______________________________________                                         Note:                                                                         The abbreviations in the table have the following significance: H             (Hydrogen), Me (Methyl group), Bu (Butyl group), Cbz (Carbobenzoxy group)     Tri (Trityl group), i- (iso-), mp (Melting point), .sup.a) (L-form).     

EXAMPLE 6

1. To a solution of carbobenzoxy-L-leucyl-glycine (4.05 g) in drymethylene chloride (50 ml), triethylamine (1.75 ml) and ethylchlorocarbonate (1.2 ml) are added at -10° C, and the mixture is stirredat the same temperature for 20 minutes. Still, a solution of2-amino-5-chlorobenzophenone (2.91 g) in dry methylene chloride (50 ml)is added at 0° C gradually thereto, and the resultant mixture is stirredunder ice cooling for 15 minutes and 1 hour and at room temperature for30 minutes and 1 hour, then refluxed overnight. The reaction mixture ispoured onto a mixture of potassium carbonate and ice and shaken withmethylene chloride. The organic layer is washed with water, dried andevaporated to remove the solvent. The residue is chromatographed on acolumn of silica gel containing water (3%), which is eluted with benzeneto recover the starting 2-amino-5-chlorobenzophenone (1.21 g) and theneluted with benzene/ethyl acetate (9:1) to give a product. The productis recrystallized from ether to give 2-benzoyl-4-chloro-N.sup.α-carbobenzoxy-L-leucylglycinanilide (3.13 g) as crystals melting at 98°to 100° C. IR: 3425, 3315, 1700, 1640 cm⁻¹ (CHCl₃).

2. In acetic acid solution (15 ml) containing hydrobromic acid (24%)2-benzoyl-4-chloro-N.sup.α -carbobenzoxy-L-leucyl-glycinanilide (3.1 g)is dissolved under ice cooling, and the resultant solution is stirred atroom temperatre for 1.5 hours. The solution is mixed with ether andallowed to stand for 30 minutes. The precipitate is filtered, dissolvedin cold water and shaken with methylene chloride/ether (1:2). Afterremoving the organic layer, the aqueous layer is made alkaline with anaqueous potassium carbonate solution, saturated with sodium chloride andshaken with chloroform. The chloroform layer is washed with water, driedover anhydrous magnesium sulfate and evaporated to remove the solvent.The residue is recrystallized from ether to give2-benzoyl-4-chloro-N.sup.α -L-leucyl-glycinanilide (1.628 g) as crystalsmelting at 145° to 147° C. IR: 3325, 1685, 1639 cm⁻¹ (CHCl₃). [α]_(D)²⁴.5 +50.7°±0.9° (EtOH). Mass, m/e 401 (M⁺).

EXAMPLES 7 to 9

Using the following starting materials (II) and (III), the reaction iseffected as in Example 6, whereby the corresponding products (Ia) and(Ib) are obtained: ##STR7## wherein R, R¹, R² and R³ are each as definedabove.

                  Table 3.                                                        ______________________________________                                        Example                                                                              II    III             Ia      Ib                                       No.    R     R.sup.1 R.sup.2                                                                             R.sup.3                                                                             mp(°(° C)                                                               mp(°  C)                      ______________________________________                                        7      H     H        Me   Cbz   131-133 143-145                              8      H     i-Pr    H     Cbz   158-168 119-121                              9      H      Ph     H     Cbz   93-95   65-67                                ______________________________________                                         Note:                                                                         The abbreviations in the table have the following significance: Pr (Propy     group), Ph (Phenyl group); the others are each as defined above.         

EXAMPLE 10

To a solution of carbobenzoxy-glycine (1.05 g) in hexamethylphosphorictriamide (8 ml), thionyl chloride (0.6 g) is added at -4° to -6° C, andthe mixture is stirred at -6° C for 10 minutes. To the mixture2-benzoyl-4-chloro-glycinanilide (1.44 g) is added, and the resultantmixture is stirred at temperature below 0° C for 2 hours and allowed tostand overnight at room temperature. The reaction mixture is madealkaline with aqueous sodium bicarbonate solution and shaken withchloroform. The organic layer is washed with water, dried and evaporatedto remove the solvent. The residue is recrystallized from ether/water togive 2-benzoyl-4-chloro-N.sup.α -carbobenzoxy-glycyl-glycinanilide (1.9g) as crystals melting at 163° to 164° C.

EXAMPLE 11

To a suspension of N,N-dimethylglycine hydrochloride (0.97 g) inanhydrous pyridine (15 ml), triphenyl phosphite (2.15 g) is added, andthe resultant mixture is stirred at room temperature overnight. Asolution of 2-benzoyl-4-chloro-N-methylglycinanilide (2.0 g) in drypyridine (10 ml) is added thereto, and the resultant mixture is stirredat room temperature for 103 hours. The reaction mixture is evaporatedunder reduced pressure. The residue is made alkaline with an aqueouspotassium carbonate solution and shaken with methylene chloride/ether (1: 2). The organic layer is washed with water and evaporated. The residueis made acidic with 3 N hydrochloric acid and shaken with ether. Afterremoving the ethereal layer, the aqueous layer is made alkaline with anaqueous potassium carbonate solution and shaken with ether. The ethereallayer is washed with a saturated saline solution, dried over anhydroussodium sulfate and evaporated to remove the solvent. The residue (1.9 g)is dissolved in methanol (2 ml), mixed with a solution of oxalic acid(0.64 g) in water (2 ml) and evaporated under reduced pressure todryness. The obtained crystals are washed with ether four times to give2-benzoyl-4-chloro-N-methyl-N.sup.α -dimethylglycyl-glycinanilideoxalate (1.75 g) as crystals melting at temperature above 90° C(decomp.). IR: 3463, 1719, 1694 (sh.), 1668 (sh.), 1640 cm⁻¹ (CHCl₃).

EXAMPLE 12

1. To a solution of N-carbobenzoxy-phenylalanine (3 g) inhexamethylphosphoric triamide (16 ml), thionyl chloride (1.2 g) is addeddropwise at -6° to -2° C in 5 minutes, and the resultant mixture isstirred at -6° to -8° C for 10 minutes. To the mixture, a suspension of1-methyl-2-aminomethyl-3-o-chlorophenyl-5-chloroindole hydrochloride(3.52 g) in ether (15 ml) previously treated with triethylamine isadded, and the resultant mixture is allowed to stand at room temperatureovernight. The reaction mixture is neutralized with an aqueous sodiumbicarbonate solution and shaken with ether. The organic layer is driedand evaporated to remove the ether. The residue is crystallized fromether to give 1-methyl-2-(N.sup.α-carbobenzoxy-phenylalanylaminomethyl)-3-o-chlorophenyl-5-chloroindole(3.15 g). This substance is recrystallized from ethyl acetate to giveneedles melting at 174° to 176° C. The yield is 54%. UV: λ_(max) ^(EtOH)232, 285 mμ (log ε=4.88, 3.28).

2. To a solution of 1-methyl-2-(N.sup.α-carbobenzoxy-phenylalanylaminomethyl)-3-o-chlorophenyl-5-chloroindole(2.86 g) in acetic acid (15 ml), a solution of chromic anhydride (1.59g) in water (1.4 ml) is added dropwise at 13° to 21° C for 5 minutes,and the resultant mixture is stirred at room temperature for 4 hours.The reaction mixture is mixed with icy water and shaken with chloroform.The organic layer is washed with water, dried and evaporated to removethe solvent. The residue is chromatographed on a column of silica gel,which is eluted with ether to give2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-carbobenzoxy-phenylalanyl-glycinanilide (1.75 g) as a gelatinoussubstance. UV: λ_(max) ^(EtOH) 256 (sh.), 298 (sh.) mμ (log ε=4.01,3.44).

3. A solution of hydrobromic acid (21.8%) in acetic acid is added to2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-carbobenzoxy-phenylalanyl-glycinanilide (1.65 g) and the resultantmixture is stirred at room temperature for 1.5 hours. The reactionmixture is mixed with dry ether, and the precipitated crystals arefiltered to give 2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-phenylalanyl-glycinanilide hydrobromide hydrate (1.3 g) as crystalsmelting at 206° to 209° C (decomp.). UV: λ_(max) ^(EtOH) 258 (sh.), 300(sh.) mμ (log ε=3.97, 3.35).

EXAMPLE 13

1. Using 1-methyl-2-aminomethyl-3-o-chlorophenyl-5-chloroindolehydrochloride and N-tritylglycine, the reaction is effected as inExample 12 (1), whereby1-methyl-2-(N-trityl-glycylaminomethyl)-3-o-chlorophenyl-5-chloroindoleis obtained as crystals melting at 198° to 200° C.

2. To a suspension of1-methyl-2-(N-trityl-glycylaminomethyl)-3-o-chlorophenyl-5-chloroindole(2.02 g) in acetic acid (10 ml), a solution of chromic anhydride (0.81g) in water (0.6 ml) is added, and the resultant mixture is stirred atroom temperature for 22 hours. The reaction mixture is mixed with water(22 ml), and the precipitated crystals are filtered. The filtrate ismixed with 28% aqueous ammonia solution (12 ml) and shaken withchloroform. The organic layer is washed with water, dried and evaporatedto remove the solvent. The residue is dissolved in ethanol and mixedwith a solution of oxalic acid in ethanol. The precipitated crystals(0.5 g) are recrystallized from ethanol to give2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α -glycyl-glycinanilideoxalate as crystals melting at temperature below 167° C. UV: λ_(max)^(EtOH) 253, 298 (sh.) mμ (log ε=3.98, 3.34).

EXAMPLE 14

1. Using 1-methyl-2-aminomethyl-3-o-chlorophenyl-5-chloroindolehydrochloride and N-carbobenzoxy-glycine, the reaction is effected as inExample 12 (1), whereby1-methyl-2-(N-carbobenzoxy-glycylaminomethyl)-3-o-chlorophenyl-5-chloroindoleis obtained as crystals melting at 96° to 98° C.

2. To a solution1-methyl-2-(N-carbobenzoxy-glycylaminomethyl)-3-o-chlorophenyl-5-chloroindole(9.1 g) in acetic acid (55 ml), a solution of chromic anhydride (5.5 g)in water (5.1 ml) is added dropwise at temperature below 20° C, and theresultant solution is allowed to stand at room temperature overnight.The reaction mixture is mixed with icy water and shaken with ethylacetate. The organic layer is washed with water, dried and evaporated toremove the solvent. The residue is chromatographed on a column of silicagel, which is eluted with ethyl acetate to give2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-carbobenzoxy-glycyl-glycinanilide (3.6 g) as a gelatinous substance.This substance is mixed with a solution of hydrobromic acid (21.8%) inacetic acid (11.5 ml) and stirred at room temperature for 1.5 hours. Thereaction mixture is mixed with ether to precipitate crystals. Thecrystals are filtered, dissolved in water and neutralized with anaqueous sodium bicarbonate solution. The precipitate is filtered to give2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α -glycyl-glycinanilidehydrate (1.8 g). This substance is recrystallized from aqueous alcoholto give prisms melting at 95°-100° C.

EXAMPLES 15 to 21

Using the following starting compounds (XI) and (V), the reactions areeffected as in Example 14, whereby the corresponding products (XII),(Ia) and (Ib) are obtained: ##STR8##

                                      Table 4.                                    __________________________________________________________________________    Example                                                                            XI   V       XII   Ia    Ib                                              No.  R  X R.sup.1                                                                          R.sup.2                                                                         R.sup.3                                                                          mp(°  C)                                                                     mp(°  C)                                                                     mp(°  C) (salt)                          __________________________________________________________________________    15.sup.a)                                                                          Me Cl                                                                              Bz H Cbz                                                                              150-155                                                                             Syrup 116˜(Oxalate)                             16.sup.b)                                                                          Me Cl                                                                              Bz H Cbz                                                                              168-169                                                                             Syrup 117-180(d) (HBr)                                17   Me Cl                                                                              H  H Cm 158-159                                                                             104-106                                                                              95-100 (H.sub.2 O)                             18   Me H i-Pr                                                                             H Cbz                                                                              242-247                                                                             Syrup ˜130 (Hemioxalate)                        19.sup.a)                                                                          Me H Bz H Cbz                                                                              255-257                                                                             Syrup 137-140 (HCl)                                   20   Me H H  H Cbz                                                                              150-153                                                                             Syrup ˜60                                       21   Me F H  H Cbz                                                                              166-167                                                                             Syrup 80˜(HCl)                                  __________________________________________________________________________     Note:                                                                         The abbreviations in this table have the following significance: Cm           (Carbomethoxy group), Cl (Chlorine), d (decomposition), F (Fluorine),         .sup.a) Levo, .sup.b) Dextro, and the others are as defined above.       

EXAMPLE 22

1. To a solution of crude1-methyl-2-aminomethyl-3-o-chlorophenyl-5-chloroindole (9.97 g) indioxane (300 ml), potassium carbonate (2.48 g) is added at roomtemperature with stirring, and the mixture is mixed withN-phthalyl-glycyl chloride (8.036 g). The resultant mixture is stirredat room temperature for 30 minutes. The reaction mixture is concentratedto a volume of about 100 ml, which is mixed with n-hexane (100 ml). Theprecipitated crystals are filtered, and dissolved in chloroform (2L)/methanol (100 ml) to give a solution, which is washed with water,dried over anhydrous sodium sulfate and evaporated to remove thesolvent. The residue is washed with ether to give 1-methyl-2-(N.sup.α-phthalyl-glycylaminomethyl)-3-o-chlorophenyl-5-chloroindole (9.642 g).The same product (450 mg) is obtained from the dioxane/n-hexane motherliquor and the ethereal washings. The yield is 62.8%. This substance isrecrystallized from methanol/chloroform to give crystals melting at 253°to 254° C.

2. To a solution of 1-methyl-2-(N.sup.α-phthalyl-glycylaminomethyl)-3-o-chlorophenyl-5-chloroindole (1.00 g) inacetic acid (25 ml), a solution of chromic anhydride (406 mg) in water(2 ml) is added gradually with stirring. The resultant mixture isstirred at 22° to 25° C for 4 hours and concentrated under reducedpressure to about half a volume. The residue is mixed with ice, and theprecipitate is filtered. The filtrate is shaken with ethyl acetate, andthe said precipitate is dissolved in the ethyl acetate layer. The ethylacetate layer is chromatographed on a column of silica gel, and theeluate is evaporated to give a precipitate, which is recrystallized frommethylene chloride/methanol to give2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-phthalyl-glycyl-glycinanilide (580 mg) as crystals melting at 216° to218° C. The yield is 54.5%.

3. To a solution of 2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-phthalyl-glycyl-glycinanilide (1.056 g) in dimethylformamide (20 ml), asolution of hydrazine hydrate (180 mg) in dimethylformamide (4 ml) isadded at -8° to -6° C with stirring, and the resultant mixture isstirred at -8° C to room temperature for 1 hour. After cooling at 0° C,the reaction mixture is mixed with N-hydrochloric acid (4 ml) in 20minutes and allowed to stand at 0° C for 17 hours. The reaction mixtureis poured into a mixture of icy water (200 ml) and ethyl acetate (100ml) and made alkaline to pH 8 with 28% aqueous ammonia solution. Theethyl acetate layer is separated, washed with water, dried overanhydrous sodium sulfate and evaporated under reduced pressure to give aresidue (500 mg). The same substance (410 mg) is obtained from theaqueous layer and washings. Both are combined, dissolved in ethanol (10ml) and mixed with water (25 ml) under cooling below 0° C. Theprecipitated crystals are filtered to give2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α -glycyl-glycinanilidehydrate (722 mg) as crystals melting at 95° to 100° C. The hemicitratemelts at 114° to 116° C. The yield is 87%.

EXAMPLES 23 TO 25

Using the following compound (XI) and N-phthalylglycyl chloride, thereactions are effected as in Example 22, whereby the correspondingproducts (XII), (Ia) and (Ib) are obtained: ##STR9##

                  Table 5.                                                        ______________________________________                                        Example                                                                              XI          XII      Ia     Ib                                         No.    R         X     mp(°  C)                                                                      mp(°  C)                                                                      mp(°  C)                          ______________________________________                                        23     Me        H     >300   --     ˜60 (Amorph)                       24     --CH.sub.2 CN                                                                           H     >300   232-234                                                                              113-120                                                                (d)    (Hemicitrate)                            25     --CH.sub.2 CN                                                                           Cl    288-289                                                                              184-186                                                                              139-143 (d)                                                     (d)           (Hemicitrate)                            ______________________________________                                         Note: The abbreviations are as defined above.                            

EXAMPLE 26

1. To a solution of 2',5-dichloro-2-methylaminobenzophenone (3.20 g) inbenzene (80 ml), phthalyl-glycylglycyl chloride (4.0 g) is added, andthe resultant mixture is stirred at 70° to 80° C for 1 hour. Theprecipitated crystals are filtered, washed with benzene and then ethanoland dried to give 2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-phthalyl-glycylglycinanilide (5.6 g), which is recrystallized fromethanol to give crystals melting at 217° C.

2. A suspension of 2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-phthalyl-glycyl-glycinanilide (81.0 g) in ethanol (50 ml) is mixed withhydrazine hydrate (20 ml), and the resultant mixture is refluxed forabout 30 minutes. After cooling, the reaction mixture is filtered toremove the insoluble phthalhydrazide. The filtrate is evaporated toremove the solvent, and the residue is crystallized from dilute ethanoland washed with ether to give2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α -glycyl-glycinanilidehydrate (55.3 g). This substance is recrystallized from dilute ethanolto give crystals melting at 95° to 100° C.

EXAMPLE 27

1. To a suspension of sodium borohydride (1.2 g) in tetrahydrofuran (10ml), a solution of 2',5-dichloro-2-methylaminobenzophenone (3.12 g) intetrahydrofuran (20 ml) is added dropwise. The resultant mixture ismixed with water (5 ml) and stirred at room temperature overnight. Thereaction mixture is mixed with a small amount of water and evaporatedunder reduced pressure to remove the solvent. The residue is made to pH8-9 with dilute hydrochloric acid and shaken with chloroform. Theorganic layer is dried and evaporated to give2',5-dichloro-2-methylaminobenzhydrol (3.05 g). This substance isrecrystallized from ether/n-hexane to give crystals melting at 105.5° to106.5° C. The yield is 97.1%.

2. To a solution of carbobenzoxy-glycyl-glycine (4.0 g) inhexamethylphosphoric triamide (20 ml)/acetonitrile (10 ml), thionylchloride (1.77 g) is added dropwise at -18° C, and the resultant mixtureis stirred for 3 minutes at -18° C. A solution of2',5-dichloro-2-methylaminobenzhydrol (2.2 g) in hexamethylphosphorictriamide (10 ml)/acetonitrile (5 ml) is added dropwise thereto at -18°C, stirred at the same temperature for 8 hours and allowed to stand at-20° C overnight. After the reaction, the reaction mixture is mixed withwater/ether, made alkaline with an aqueous sodium bicarbonate solutionand shaken with ether. The organic layer is dried and evaporated toremove the solvent. The residue is chromatographed on a column of silicagel, which is eluted with ethyl acetate to give2-o-chloro-α-hydroxybenzyl-4-chloro-N-methyl-N.sup.α-carbobenzoxy-glycyl-glycinanilide (3.31 g) as crystals melting at 57°to 60° C.

3. To a solution of 2-o-chloro-α-hydroxybenzyl-N-methyl-N.sup.α-carbobenzoxy-glycyl-glycinanilide (21.8 g) in acetone (300 ml), Jonesreagent (a solution of chromic acid and sulfuric acid in water) is addeddropwise until the reaction mixture keeps red. The reaction mixture isfiltered to remove the precipitate. The red filtrate is mixed withisopropanol until the red solution becomes green. The mixture isfiltered, and the filtrate is neutralized with an aqueous sodiumbicarbonate solution and evaporated. The residue is mixed with water andshaken with chloroform. The chloroform layer is purified with activecarbon to give 2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-carbobenzoxy-glycyl-glycinanilide (21.3 g). The yield is 98.5%.

EXAMPLES 28 to 33

Using the following compound (IX), the reactions are effected as inExample 27 but when phthalyl group is adopted for amino-protection, itis removed by hydrazinolysis, whereby the following compounds (X), (Ia)and (Ib) are obtained: ##STR10##

                  Table 6.                                                        ______________________________________                                        Ex.  IX       X                Ia     Ib                                      No.  R      X     R.sup.1                                                                            R.sup.2                                                                           R.sup.3                                                                            mp(°  C)                                                                      mp(°  C)                                                                      mp(°                     ______________________________________                                                                                      C)                              28   Me     H     H    H   Cbz  75-78  45-50  ˜60 (A-                                                                 morph)                          29.sup.a)                                                                          Me     Cl    Bz   H   Cbz  70     Amorph 110˜(A-                                                                 morph)                          30   Me     Cl    H    Ft     200-201                                                                              223-226                                                                              95-100                                                                        (H.sub.2)0                        31   Me     F     H    Ft     193-194                                                                              213-214                                                                               80˜                                                      (d)           (HCl)                             32   De     F     H    Ft     166-168                                                                              186-187                                                                              Amorph                                                          (d)                                             33   Me     Cl    i-Pr H   Cbz  172-173                                                                              --     100                                                                           (HCl)                           ______________________________________                                         Note:                                                                         The abbreviations have the following significance: Ft (Phthalyl group), D     (Diethylaminoethyl group), .sup.a)  Levo, and the others are as defined       above.                                                                   

EXAMPLE 34

1. To a solution of 2-o-chlorobenzoyl-4-chloro-N-methyl-glycinanilidehydrobromide (1.8 g) in hexamethyl-phosphoric triamide (10 ml),chloroacetyl chloride (0.73 g) is added under ice cooling. The resultantmixture is stirred under ice cooling for 2 hours and at room temperaturefor 3 hours. The reaction mixture is shaken with ether and the organiclayer is made alkaline with an aqueous ammonia solution, washed withwater and evaporated to give 2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-chloroacetyl-glycinanilide (1.6 g). This substance is recrystallizedfrom ethyl acetate to give colorless needles melting at 134° to 136° C.

2. A mixture of 2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-chloroacetyl-glycinanilide (6.2 g), potassium iodide (2.74 g) andacetone (60 ml) is refluxed for 1 hour. The reaction mixture isevaporated to remove the acetone, and the residue is dissolved inchloroform. The organic layer is washed with water, dried andevaporated. The residue is washed with ether to give2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α -iodoacetyl-glycinanilide(6.9 g). This substance is recrystallized from ethyl acetate to givecolorless needles melting at 168.5 to 169.5° C.

3. Into a suspension of 2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-iodoacetyl-glycinanilide (1.1 g) in tetrahydrofuran (20 ml) ammonia gasis introduced for 30 minutes, and the mixture is stirred at roomtemperature for 5 hours. The reaction mixture is evaporated to removethe tetrahydrofuran. The residue is dissolved in chloroform. The organiclayer is washed with an aqueous sodium bicarbonate solution and thenwater, dried and evaporated to remove the chloroform. The residue ischromatographed on a column of silica gel, which is eluted with methanolto give 2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-glycyl-glycinanilide.

EXAMPLE 35

1. The reaction is effected as in Example 34 (1) by using bromoacetylbromide in liue of chloroacetyl chloride, whereby2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α -bromoacetyl-glycinanilideis obtained as colorless needles melting at 153° to 155° C. The yield is69%.

2. To a solution of 2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-bromoacetyl-glycinanilide (1.01 g) in dimethylformamide (10 ml),potassium phthalimide (0.34 g) is added, and the resultant mixture isstirred at room temperature for 3 hours and allowed to stand at roomtemperature overnight. The reaction mixture is mixed with water (100ml), and the precipitate is filtered and washed with water to give2-o-chloro-benzoyl-4-chloro-N-methyl-N.sup.α-phthalyl-glycyl-glycinanilide (1.0 g).

EXAMPLE 36

1. To a solution of 2-o-chlorobenzoyl-4-chloro-N-methyl-glycinanilidehydrobromide (0.628 g) in dimethylformamide (7 ml), phthalyl-glycylchloride (0.437 g) is added, and the resultant mixture is stirred for 3hours. The reaction mixture is evaporated to remove the solvent. Theresidue is shaken with chloroform, and the chloroform layer is washedwith water, dried and evaporated. The residue is washed with ether togive 2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-phthalyl-glycyl-glycinanilide (0.71 g). The yield is 93.5%.

2. The above product is treated with hydrazine hydrate to give2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α -glycyl-glycinanilidehydrate.

EXAMPLES 37 TO 46

Using the following amines in lieu of ammonia, the reactions areeffected as in Example 34 (3), whereby the corresponding products (I)are obtained: ##STR11##

                  Table 7.                                                        ______________________________________                                        Example   VIII              I                                                 No.     R.sup.2 --NH--R.sup.3                                                                           mp(°  C) (Salt)                              ______________________________________                                        37      Piperidine         81-83                                              38      Diethylamine      113-115                                             39      4-hydroxy-4-(p-chlorophe-                                                     nyl)piperidine    144-146                                             40      Dimethylamine     132-133                                             41      Morpholine        120-122                                             42      Methylamine       100-102                                             43      4-phenylpiperazine                                                                              155-164 (d) (2HCl)                                  44      4-methylpiperazine                                                                              226-228 (d) (2HCl)                                  45      isopropylamine    197-200 (d) (HCl)                                   46      Phenethylamine    176-178 (Oxalate)                                   ______________________________________                                         Note: The abbreviations are as defined above.                            

EXAMPLES 47 TO 50

The reactions are effected as in Example 1, whereby the followingproducts (I) are obtained:

    ______________________________________                                        Example                                                                       No.    Compound Name           mp(°  C)                                ______________________________________                                        47     2-(α-picolyl)-4-chloro-N.sup.α -glycyl-glycin-                                            192-194 (d)                                           anilide dihydrobromide                                                 48     3-benzoyl-5-ethyl-2-(N.sup.α -glycyl-                                                           190-192                                               glycyl)amino-thiophene hydrochloride                                   49     2-o-chlorobenzoyl-4-chloro-N-methyl-                                                                  100                                                   N.sup.α -diglycyl-glycinanilide                                  50     2-o-chlorobenzoyl-4-chloro-N-methyl-                                                                  Amorph                                                N.sup.60 -triglycyl-glycinanilide                                      ______________________________________                                    

What we claim is:
 1. A process for preparing2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α -glycyl-glycinanilide whichcomprises reacting1-methyl-2-aminomethyl-3-o-chlorophenyl-5-chloroindole withphthalyl-glycyl chloride in an inert solvent, oxidizing the resultant1-methyl-2-(N.sup.α-phthalyl-glycylaminomethyl)-3-o-chlorophenyl-5-chloroindole withchromic anhydride/acetic acid and subjecting the resultant2-o-chlorobenzoyl-4-chloro-N-methyl-N.sup.α-phthalylglycyl-glycinanilide to hydrazinolysis in an inert solvent.